AlphaScreen SureFire IKKa (p-Ser176/Ser180) Assay Kit - 10,000 Points

The AlphaScreen®SureFire® p-IKKα (Ser176/180) assay is a sandwich immunoassay for quantitative detection of phospho-IKKα (phosphorylated on Ser176/180) in cellular lysates using Alpha Technology. Please note that an AlphaScreen Protein A kit, which contains the Alpha Beads necessary to run this assay, needs to be purchased separately in addition to the AlphaScreen SureFire kit.

The HV 96-well kits contain enough reagents to run 100 wells in 96-well format, using a 50 µL reaction volume.

The 500 point kit contains enough reagents to run 500 wells in 384-well format, using an 11 µL reaction volume.

The 10,000 point kit contains reagents to run 10,000 wells in 384-well format, using an 11 µL reaction volume.

The 50,000 point kit contains enough reagents to run 50,000 wells in 384-well format, using an 11 µL reaction volume.

In the AlphaScreen® SureFire® assay, Donor beads are coated with streptavidin to capture one of the antibodies, which is biotinylated. Acceptor beads are coated with Protein A to immobilize the other antibody. In the presence of phosphorylated protein, the two antibodies bring the Donor and Acceptor beads close together, generating signal. The amount of light emission is directly proportional to the amount of phosphoprotein present in the sample.

Screening kinase activities in cell-based assays offers advantages over the more traditional biochemical approach of using a purified recombinant enzyme to phosphorylate a substrate, since a cell-based assay gives information on a compound’s activity in a more biological context. However, getting initial results with the AlphaScreen® SureFire® assay is highly dependent on optimal cell culture conditions, and often requires that multiple parametersbe optimized in the first set of experiments.